Within the last several years, a number of ependymins have been molecularly cloned from a variety of teleost fish including Oncorhynchus mykiss (rainbow trout; Muller-Schmid, A., et al., Gene 118:189-196 (1992)), Salmo salar (Atlantic salmon; Muller-Schmid, A., et al., Gene 118:189-196 (1992)), Esox lucius (pike; Muller-Schmid, A., et al., J. Molec. Evol. 36:578-585 (1993)), Carassius auratus (goldfish; Konigstorfer, A., et al., J. Neurochem. 52:310-312 (1989); Konigstorfer, A., et al., J. Biol. Chem. 264:13689-13692 (1989)), Brachydanio rerio (zebrafish; Sterrer, S., et al., Neurosci. 37:277-284 (1990)), and Clupea harengus (herring, Muller-Schmid, et al., J. Molec. Evol. 36:578-585 (1993)). The ependymins produced by these organisms are synthesized as precursors which contain N-terminal, hydrophobic signal sequences. Each of these molecules contains multiple N-linked glycosylation sites, only some of which are conserved between species (Schmidt, R. and Shashoua, V. E. J. Neurochem. 36:1368-1377 (1981); Schmidt, R. and Shashoua, V. E. J. Neurochem. 40:652-660 (1983); Ganb, B. and Hoffman, W. Eur. J. Biochem. 217:275-280 (1993)). The precursor ependymins range in apparent molecular mass from 23.7 to 24.5 kDa, while the secreted mature forms of these molecules are typically 21.6-22.3 kDa in size.
The piscine ependymins characterized thus far may be categorized according to the number of cysteine residues present in the mature polypeptide. Mature salmoniform (O. mykiss, S. salar, and E. lucius) ependymin polypeptides contain only four cysteine residues, whereas mature cypriniform (C. auratus and B. rerio) and clupeiform (C. harengus) ependymin polypeptides contain five and six cysteine residues, respectively (Hoffmann, W. Int. J. Biochem. 26:607-619 (1994)). Correspondingly, disulfide-linked dimerization of the salmoniform ependymin polypeptides is not observed after non-reducing SDS-PAGE. However, cypriniform and clupeiform ependymins are observed as disulfide-linked dimers under non-reducing conditions. It is speculated that the dimerization occurs via the cysteine residue conserved only between the salmoniform ependymins (this cysteine residue aligns with the lysine residue at location 133 of human ependymin of the present invention as shown in SEQ ID NO:2).
Several lines of evidence have provided the basis for an understanding of the functional role(s) of the ependymins. Ca2+-binding has been demonstrated for at least goldfish and rainbow trout ependymins (Schmidt, R. and Makiola, E. Neuro. Chem. (Life Sci. Adv.) 10:161-171 (1991); Ganb, B. and Hoffman, supra). Further, ependymins are the primary cerebrospinal fluid component in a number of teleost fish (Schmidt, R. and Lapp, H. Neurochem. Int. 10:383-390 1987). Finally, roughly two-thirds of Ca2+ in the CSF of rainbow trout is protein-bound (Ganb, B. and Hoffman, supra). As a result, it is thought that ependymins may function in Ca2+ homeostasis of the teleost piscine brain (Hoffman, W., supra).
In situ hybridization analyses have shown that ependymins are apparently synthesized exclusively in miningeal fibroblasts of the mininx (also termed the endomeninx of leptomeninx) of teleost fish (Konigstorfer, A., et al., Cell Tissue Res. 261:59-64 (1990)). Ependymins have also been found to associate with collagen fibrils of the extracellular matrix (ECM; Schwarz, H., et al. Cell Tissue Res. 273:417-425 (1993)), and, further, have the capacity to serve as a substrate for outgrowing retinal axons (Schmidt, J. T., et al., J. Neurobiol. 22:40-54 (1991)).
An additional role for ependymins has been identified in the field of learning and memory. Using an experimental approach in which goldfish learn to swim to a specific compartment of its environment to avoid an electric shock, investigators have determined that the amount of unbound or unincorporated extracellular ependymins decreases after learning (Piront, M.-L., and Schmidt, R. Brain Res. 442:53-62 (1988); Schmidt, R. J. Neurochem. 48:1870-1878 (1987)). Further, blockage of functional ependymin molecules, either with antibodies or antisense polynucleotides, resulted in the reversible inability of the experimental animal to remember the task which it had learned. Removal of the inhibitory substance then resulted in a reappearance of the learned ability (Schmidt, R. J. supra; Shashoua, V. E. and Moore, M. E. Brain Res. 148:441-449 (1978)).
Thus, there is a need for polypeptides that function as neurotrophic factors in the regeneration of the optic and other nerves and in long-term memory consolidation, since disturbances of such regulation may be involved in disorders relating to the complex molecular and cellular process regulating neuronal and nervous system function. Such disorders may include Parkinson's disease, Alzheimer's disease, amyotrophic lateral sclerosis, pain, stroke, depression, anxiety, epilepsy, and other neurological and psychiatric disorders. Therefore, there is a need for identification and characterization of such human polypeptides which can play a role in detecting, preventing, ameliorating or correcting such disorders.